EMT6 is a BALB/c mammary tumor which is quite immunogenic in syngeneic mice. Despite the ability of the immune system to recognize this tumor as foreign and to generate specific cytolytic T cells, this tumor nevertheless grows progressively resulting in the death of the mice. We have attempted to expand the specifically reactive T cell population in vitro by incubating cells with IL-2 and antigen in the form of inactivated (irradiated) tumor cells. However, it was found that even small numbers of irradiated tumor cells were very inhibitory to the growth of the T cells. Further investigations have shown that EMT6 tumor cells produce an immunoinhibitory factor (IIF) which prevents the proliferation of lymphocytes. We have been unable to find any form of the tumor cell antigen which is stimulatory. Since propagation of the tumor reactive T cells requires stimulation of the antigen specific receptor, we are exploring the possibility of stimulating the cells nonspecifically using a hamster monoclonal antibody reactive with CD3, an invariant portion of the T cell receptor complex. Specifically reactive T cells have been preselected by removing them from growing tumors and then stimulated. Three cell lines have been obtained and are currently being analyzed for function and phenotype. These cell lines will be expanded and used for immunotherapy protocols in vivo.